Throughout this application various publications are referred to by partial citations within parentheses. Full citations for these publications may be found at the end of the specification immediately preceding the claims. The disclosures of these publications, in their entireties, are hereby incorporated by reference into this application in order to describe more fully the state of the art to which the invention pertains.
Neuroregulators comprise a diverse group of natural products that subverse or modulate communication in the nervous system. They include, but are not limited to, neuropeptides, amino acids, biogenic amines, lipids, and lipid metabolites, and other metabolic byproducts. Many of these neuroregulator substances interact with specific cell surface receptors, which transduce signals from the outside to the inside of the cell. G-protein coupled receptors (GPCRs) represent a major class of cell surface receptors with which many neurotransmitters interact to mediate their effects. GPCRs are characterized by seven membrane-spanning domains and are coupled to their effectors via G-proteins linking receptor activation with intracellular biochemical sequelae such as stimulation of adenylyl cyclase.
Tyramine (TYR), β-phenyl-ethylamine (PEA), tryptamine (T) and octopamine (OCT) belong to a class of amines that have low endogenous levels in tissues and thus are referred to as “trace amines” (Usdin and Sandler, 1976). For example, under physiological conditions, brain levels of T in the rat are a thousand-fold lower than those of 5-hydroxytryptamine (5-HT), a major neurotransmitter in vertebrates and invertebrates (Artigas and Gelpi, 1979).
In invertebrates, the role of “trace amines” as neurotransmitters is well established, in particular for OCT and TYR, whose physiological actions have been shown to be mediated directly via their specific receptors. Octopamine, the monohydroxylated analogue of NE, has been studied the most in this respect and is a major neurotransmitter, neurohormone and neuromodulator in many invertebrate species (Axelrod and Saavedra, 1977; David and Coulon, 1985). Because many of the octopamine-mediated responses are connected to adaptation to stressful circumstances, the octopaminergic system has been considered to be the invertebrate equivalent of the vertebrate sympathetic nervous system. Recently, the cloning of the first invertebrate (mollusc) OCT receptor has been reported and it belongs to the family of G protein coupled receptors (GPCR) (Gerhardt et al., 1997). Similarly, TYR which is the precursor of OCT, is abundant in insect brains and its distribution in different tissues does not parallel that of OCT, suggesting that TYR is not merely a precursor in the biosynthetic pathway of OCT (Juorio and Sloley, 1988; Maxwell et al., 1978). In fact, TYR and OCT have opposite effects on adenylate cyclase and glycogenolysis in cockroach fat bodies, TYR being inhibitory and OCT being stimulatory (Downer, 1979). Therefore, in addition to OCT, TYR has also been suggested to play a role as a neurotransmitter in invertebrates (Roeder, 1994). Cloning of an adenylate cyclase inhibitory Drosophila TYR receptor, belonging to the family of GPCRs (Saudou et al., 1990) supports this hypothesis.
The evidence for the role of “trace amines” as neurotransmitters in the mammalian system has not been carefully studied. Because of the low concentrations (<100 ng/g) of “trace amines” in mammalian tissues it has sometimes been suggested that they might occur as by-products in the synthesis of other amine neurotransmitters such as the catecholamines or 5-HT. It is now apparent that the turnover of the “trace amines” in most tissues is very rapid, as evidenced by their loss from the brain after intraventricular administration (Wu and Boulton, 1973) and by their accumulation after inhibition of their major catabolic enzyme, monoamine oxidase (MAO) (Axelrod and Saavedra, 1974; Juorio and Durden, 1984; Philips and Boulton, 1979). Due to the fact that these “trace amines” share synthetic and catabolic enzymes with the classical amines, such as 5-HT, norepinephrine (NE) and dopamine (DA), they have also been referred to as “false transmitters” (McGeer et al., 1979). These amines are thus taken up by aminergic neurons, displace monoamines from their storage sites in vesicles, and can themselves and/or other amine neurotransmitters, then be released from neurons by electrical stimulation. Therefore, in mammals, some of the physiological actions of these “trace amines” (sympathomimetic in general, pressor, cardiac stimulant and vasoconstrictor activity) are primarily indirect and are caused by a release of endogenous neurotransmitters (NE, 5-HT, DA).
However, there is a growing body of evidence suggesting that “trace amines” function independently of the classical amine neurotransmitters and mediate some of their effects via their specific receptors. Some of these are described below.
Tyramine is among the first “trace amines” subjected to experimental study. Radiolabeled TYR can be released from rat striatal slices following KCl− depolarization. In reserpine pretreated rats, TYR induced a marked increase in the motor activity, which was not accompanied by a significant decrease in brain catecholamines, ruling out the possibility of indirect receptor stimulation (Stoof et al., 1976). A direct endothelium- and β2-adrenoceptor independent vasorelaxant effect of TYR on rat aortic strips has been reported (Varma and Chemtob, 1993; Varma et al., 1995). Saturable binding sites for [3H]p-tyramine have been reported in rat brain, which may reflect specific TYR receptor sites (Ungar et al., 1977; Vaccaria, 1986; Vaccaria, 1988). Further studies are needed before a clear definition of specific p-tyramine binding site is available. There are no reports of m-tyramine binding sites available as yet.
β-Phenylethylamine which has a chemical structure and pharmacological and behavioral effects that closely resemble those of amphetamine (evokes stereotyped behavior, anorexia and increases locomotor activity) (Wolf and Mosnaim, 1983) and has been described as the body's natural amphetamine. β-Phenylethylamine is synthesized in and released by dopaminergic neurons of the nigrostriatal system (Greenshaw et al., 1986). Saturable, high affinity binding sites have been reported for [3H]β-Phenylethylamine (Hauger et al., 1982). The highest concentration of binding sites was in the hypothalamus, where highest endogenous levels of this amine has been reported (Durden and Philips, 1973). Interestingly, saturable binding sites have also been reported for [3H]amphetamine in membrane preparations from rat brain (Paul et al., 1982), the density of these binding sites being highest in the hypothalamus, as has been seen with PEA binding. These binding sites were shown not to be associated with any previously described neurotransmitter or drug receptor sites and were specific to amphetamine and related PEA derivatives. Furthermore, the relative affinities of a series of PEA derivatives for this binding site were highly correlated to their potencies as anorexic agents. These results suggest the presence of specific receptor sites in the hypothalamus that mediate the anorexic activity of amphetamine and related PEAs.
In addition to TYR and PEA, T has also been shown to produce several physiological effects that are direct and distinct from those mediated by other aminergic neurotransmitters. Tryptamine has been shown to have opposite effects to 5-HT in several Systems studied. For example, unilateral intrahypothalamic injection of T into the preoptic area of the rat causes hyperthermia, whereas 5-HT administered into the same area produces the opposite effect (Cox et al., 1981; Cox et al., 1983). Intravenous administration of T to young rats leads to behavioral stimulation and electrocortical desynchronization, whereas behavioral depression and electrical synchronization was evoked by 5-HT (Dewhurst and Marley, 1965). Also, iontophoretic application of 5-HT and T to cortical neurons has been noted to produce excitatory and inhibitory responses, respectively (Jones, 1982b,c). Injection of deuterated T into the nucleus accumbens of the rat produces sustained locomotor stimulation (Marien et al., 1987), whereas 5-HT injection into the same area produces either only a transient decrease in locomotor activity (Pijnenburg et al., 1976) or no significant effect on locomotion (Gerber et al., 1986; Jackson et al., 1975; Kitada et al., 1983; Plaznik et al., 1985). Tryptamine as well as 5-HT cause contraction of the rat stomach fundus. However, using the non-selective antagonist, phenoxybenzamine (PBZ), Winter and Gessner (1968) showed that the T-induced contractions were more resistant to PBZ blockade than 5-HT-induced contractions. Also, tetrahydro-β-carboline (THBC) antagonizes tryptamine, but not 5-HT-mediated contraction of the isolated rat tail artery (Hicks and Langer, 1983).
The presence of specific, saturable and high affinity [3H]-T binding sites in the rat brain (Altar et al., 1986; Kellar and Cascio, 1982; McCormack et al., 1986; Perry, 1986) and peripheral tissue (Brüning and Rommelspacher, 1984) has been known for a few years. The pharmacological profile of [3H]-T binding is distinct and does not correspond to any known neurotransmitter, transporter or MAO site (Biegon et al., 1982; Fuxe et al., 1983; Leysen et al., 1982; Meibach et al., 1980, 1982; Nakada et al., 1984; Palacios et al., 1983; Perry, 1986, 1988; Slater and Patel, 1983).
The existence of p-octopamine binding sites has been demonstrated in crude membranes obtained from fruitflies but not shown so far in vertebrates (Dudai, 1982; Dudai and Zvi, 1984; Hashemzadeh et al., 1985).
The above findings indicate that in the mammalian system, TYR, PEA and T may function as neurotransmitters in their own rights, and mediate their responses via acting at their distinctive receptors.
“Trace amines” could play a role in depression and psychiatric disorders as well as migraine. Clinical literature supports these indications. MAO inhibitors that are clinically effective for the treatment of depression in the human have been shown to produce a proportionally greater increase in “trace amine” levels compared to 5-HT levels (Boulton, 1976; Juorio, 1976). Based on a functional deficiency of “trace amines”, PEA and/or T in particular, have been proposed as a possible etiological factor in depression in humans (Dewhurst, 1968a, b; Dewhurst and Marley, 1965; Sabelli and Monsnaim, 1974). The urinary output of T has been shown to be disturbed in schizophrenic patients (Brune and Himwhich, 1962; Herkert and Keup, 1969) and in the general psychiatric population (Slingsby and Boulton, 1976). The urinary output of T seems to be positively correlated with increasing severity of psychosis (Brune and Himwhich, 1962; Herkert and Keup, 1969). Depressed patients on the other hand, exhibit decreased urinary output of T (Coppen et al., 1965) and OCT (Sandler et al., 1979).
A role of “trace amines” in migraine is implicated, since certain pharmacological agents in food, in particular TYR, are believed to provoke migraine. There are many reports that attacks of palpitation, hypertension and severe headache (the so called “cheese effect”) may follow the consumption of food containing TYR in patients being treated with MAO inhibitors (see Vaughan, 1994 for review). Furthermore, clinical studies have shown that migraine sufferers had lower urinary excretion of TYR sulphate following oral TYR challenge than normal controls. The lower TYR sulfate excretion values among patients with both migraine and depression compared to those of migraine alone or depression alone suggest that comorbid migraine with depression may represent a more severe form of migraine than migraine alone (Merikangas et al., 1995). It is likely that disturbances in the same neurochemical systems, most probably the “trace amines”, account for the co-occurrence of migraine and depression.
Urinary levels of PEA, TYR and indole-3-acetic acid (the acid metabolite of T) were found to be decreased in Tourette's Syndrome (TS) patients when compared to values in normal children, indicating a role of these “trace amines” in TS (Baker et al., 1993). Urinary levels of PEA have been shown to be significantly lower in patients with learning disability (LD) and in patients suffering from Attention Deficit Hyperactivity Disorder (ADHD) as compared to age-matched controls, indicating an important role of PEA in pathogenesis of LD and ADHD (Matsuishi and Yamashita, 1999). Tryptamine has also been implicated to play a role in Parkinson's disease, since Parkinsonian patients excrete abnormally high levels of T in their urine (Smith and Kellow, 1969).
Altered “trace amine” metabolism has been observed in non-psychiatric conditions such as pellagra (Sullivan, 1922), Hartnup's disease (Baron et al., 1956), phenylketonuria (Armstrong and Robinson, 1954; Perry, 1962) and thyrotoxicosis (Levine et al., 1962).
Studies in non-human species, rats and mice in particular, add further support for some of the roles of the “trace amines” described above as well as providing various additional physiological roles of “trace amines”, as discussed below.
Interestingly, MAO A knock-out mice have elevated brain levels of 5-HT, NE and DA and manifest aggressive behavior similar to human males with a deletion of MAO A. In contrast, MAO B knock-out mice do not exhibit aggression and only levels of PEA are increased. Mice lacking MAO B are resistant to the Parkinsongenic neurotoxin, 1-methyl-4-phenyl-1,2,3,6-tetra-hydropyridine (MPP+) (Shih et al., 1999), indicating that PEA may provide neuroprotection. Both MAO A and MAO B knock-out mice show increased reactivity to stress suggesting a role of PEA in this condition.
A possible role for T in the protection against renal hypertension afforded by TRP has been suggested (Fregly et al., 1988). A role of OCT in hypertension has been suggested since a hypertensive strain of rats (SHR Kyoto) demonstrates considerably elevated levels of this amine in their brain (David, 1978; 1979). Housing stress has been shown in rats to cause an increase brain and adrenal T levels (Harrison and Christian, 1984) which may be the cause of cardiovascular changes (Bennett and Gardiner, 1978) and hyperactivity (Weinstock et al., 1978) observed in these animals. Therefore, T has been proposed to play a role in is the physiological, behavioral and chemical response to psychological stress.
Tryptamine's actions in the stomach and the presence of [3H]-T binding sites in the stomach suggest a role for T in gastric emptying and control of secretory processes (Brüning and Rommelspacher, 1984; Cohen and Wittenauer, 1985; Winter and Gessner, 1968).
Tryptamine has also been suggested to play a role in hepatic encephalopathy where, due to liver failure, there is a massive increase in brain TRP (precursor of T) leading to a series of CNS symptoms including altered sleep patterns and personality changes and eventually resulting in coma (Sourkes, 1978).
Tryptamine has been shown to cause release from isolated rat lungs of a spasmogen, resembling slow reacting substance of anaphylaxis that has prostaglandin E-like activity (Bakhle and Smith, 1977). Therefore, T may have a function in asthma.
In summary, “trace amines” may act as neurotransmitters and neuromodulators. These amines may act via their specific receptor sites to elicit some of their physiological actions. It is not yet clear what the role of these “trace amines” is in pathological conditions such as mental and physical stress, hepatic dysfunction, hypertension and electrolyte imbalance. A primary role of “trace amines” in the etiology of mental or neurological diseases is still hypothetical. “Trace amine”-mediated effects indicate that receptors for these amines are attractive as targets for therapeutic intervention for several disorders and would be useful to develop drugs with higher specificity and fewer side effects for a wide variety of diseases.